The ongoing purpose of this project is to study the genome of the Aleutian mink disease parvovirus (ADV) and to relate structural features of the genome to functional correlates, such as antigenic epitopes, pathogenicity determinants, strain variation and host range. By constructing additional full-length chimeric molecular clones between the cell-culture adapted ADV-G and the pathogenic ADV-Utah isolates, we have developed a virus, XXX-J-8-10, that is replication competent for cell culture and also is infectious for mink. Animals injected with this virus developed typical pathological findings of AD. XXX-J-8-10 is entirely ADV-G except for a portion of the capsid protein sequence (73-88 map units) which is ADV-Utah. This pathogenic and nonpathogenic viral isolates differ only by 4 amino acid residues in this region. This portion of the capsid protein sequence contains the analogue to surface loop 4 of canine parvovirus (CPV), a region implicated in host range and pathogenicity of that virus. Other chimeras prepared during the development of XXX-J-8-10 revealed that there are at least 2 distinct determinants in the capsid protein sequence that govern replication competence in vitro. The capsid protein gene of ADV was expressed in 10 non-overlapping segments in a prokaryotic expression vector, pMAL-c2. Sera from infected mink exhibited preferential reactivity for selected regions. The areas corresponding to the VP1 unique sequence and CPV surface loops 3 and 4 were consistently reactive. The hypervariable region of the capsid protein, corresponding to CPV surface loop 2, was found to contain a type specific linear epitope recognized by heterologous antisera, a monoclonal antibody and mink sera. Heterologous antisera were prepared against the analogues of the CPV surface loops and the VP1 unique regions. Although all reacted in immunoblot against the bona fide ADV capsid proteins(s), only the antisera against loops 3 and 4 were able to neutralize ADV infectivity in vitro and bind to purified capsids in immune electron microscopy. These findings define a pivotal role for the surface loops of the ADV capsid proteins. Computer analysis suggests that loops 3 and 4 form the spike located at the 3 fold axes of symmetry on the viral particle. Studies to derive the structure of the ADV virion by cryo-electron microscopy and x-ray crystallography have been initiated using empty capsids expressed in a recombinant baculovirus.